1. Field of the Invention
The present invention relates generally to electrochemical sensors that can be used for the quantification of a specific component or analyte in a liquid sample. Particularly, this invention relates to a new and improved electrochemical sensor and to a new and improved method of fabricating electrochemical sensors. More particularly, this invention relates to a disposable electrochemical sensor that is inexpensive to manufacture. Even more particularly, this invention relates to a disposable electrochemical sensor that gives accurate readings in the presence of interferants and varying red blood cells (hematocrit). Still even more particularly, this invention relates to disposable electrochemical sensors which are used for performing electrochemical assays for the accurate determination of analytes in physiological fluids.
2. Description of the Prior Art
Biosensors have been known for more than three decades. They are used to determine concentrations of various analytes in fluids. Of particular interest is the measurement of blood glucose. It is well known that the concentration of blood glucose is extremely important for maintaining homeostasis. Products that measure fluctuations in a person's blood sugar, or glucose levels have become everyday necessities for many of the nation's millions of diabetics. Because this disorder can cause dangerous anomalies in blood chemistry and is believed to be a contributor to vision loss and kidney failure, most diabetics need to test themselves periodically and adjust their glucose level accordingly, usually with insulin injections. If the concentration of blood glucose is below the normal range, patients can suffer from unconsciousness and lowered blood pressure which may even result in death. If the blood glucose concentration is higher than the normal range, the excess blood glucose can result in synthesis of fatty acids and cholesterol, and in diabetics, coma. Thus, the measurement of blood glucose levels has become a daily necessity for diabetic individuals who control their level of blood glucose by insulin therapy.
Patients who are insulin dependent are instructed by doctors to check their blood-sugar levels as often as four times a day. To accommodate a normal life style to the need of frequent monitoring of glucose levels, home blood glucose testing was made available with the development of reagent strips for whole blood testing.
One type of blood glucose biosensors is an enzyme electrode combined with a mediator compound which shuttles electrons between the enzyme and the electrode resulting in a measurable current signal when glucose is present. The most commonly used mediators are potassium ferricyanide, ferrocene and its derivatives, as well as other metal-complexes. Many sensors based on this second type of electrode have been disclosed. Examples of this type of device are disclosed in the following patents.
U.S. Pat. No. 5,628,890 (1997, Carter et al.) Disclosed an electrode strip having an electrode support, a reference or counter electrode disposed on the support, a working electrode spaced from the reference or counter electrode on the support, a covering layer defining an enclosed space over the reference and working electrodes and having an aperture for receiving a sample into the enclosed space, and a plurality of mesh layers interposed in the enclosed space between the covering layer and the support. The covering layer has a sample application aperture spaced from the electrodes. The working electrode includes an enzyme capable of catalyzing a reaction involving a substrate for the enzyme and a mediator capable of transferring electrons between the enzyme-catalyzed reaction and the working electrode.
This device proposes to reduce the effect of hematocrit on the sensor readings. According to the disclosure, this results from the downstream spacing of the reference electrode relative to the working electrode in combination with the thin layer of the sample solution created by the mesh layers.
U.S. Pat. No. 5,708,247 (1998, McAleer et al.) Disclosed a disposable glucose test strip having a substrate, a reference electrode, a working electrode, and a means for making an electrical connection. The working electrode has a conductive base layer and a coating layer disposed over the conductive base layer. The coating layer is a filler having both hydrophobic and hydrophilic surface regions which form a network, an enzyme and a mediator.
U.S. Pat. No. 5,682,884 (1997, Hill et al.) Disclosed a strip electrode with screen printing. The strip has an elongated support which includes a first and second conductor each extending along the support. An active electrode, positioned to contact the liquid mixture and the first conductor, has a deposit of an enzyme capable of catalyzing a reaction and an electron mediator. A reference electrode is positioned to contact the mixture and the second conductor.
U.S. Pat. No. 5,759,364 (1998, Charlton et al.) Disclosed an electrochemical biosensor having an insulating base plate bearing an electrode on its surface which reacts with an analyte to produce mobile electrons. The base plate is mated with a lid of deformable material which has a concave area surrounded by a flat surface so that when mated to the base plate there is formed a capillary space into which a fluid test sample can be drawn. The side of the lid facing the base is coated with a polymeric material which serves to bond the lid to the base plate and to increase the hydrophilic nature of the capillary space.
U.S. Pat. No. 5,762,770 (1998, Pritchard et al.) Disclosed an electrochemical biosensor test strip that has a minimum volume blood sample requirement of about 9 microliters. The test strip has a working and counter electrodes that are substantially the same size and made of the same electrically conducting material placed on a first insulating substrate. Overlaying the electrodes is a second insulating substrate which includes a cutout portion that forms a reagent well. The cutout portion exposes a smaller area of the counter electrode than the working electrode. A reagent for analysis of an analyte substantially covers the exposed areas of the working and counter electrodes in the reagent well. Overlaying the reagent well and affixed to the second insulating substrate is a spreading mesh that is impregnated with a surfactant.
U.S. Pat. No. 5,755,953 (1998, Henning et al.) Disclosed an reduced-interference biosensor. The device generally comprises an electrode used to electrochemically measure the concentration of an analyte of interest in a solution. The device includes a peroxidase enzyme covalently bound to microparticle carbon and retained in a matrix in intimate contact with the electrode. According to this disclosure, it is the enzyme/microparticle carbon of the device which provides a composition which is displays little sensitivity to known interfering substances.
U.S. Pat. No. 5,120,420 (1992, Nankai et al.) Disclosed a biosensor with a base board having an electrode system mainly made of carbon, an insulating layer, a reaction layer containing an enzyme layer thereon, a spacer and a cover. The spacer creates a channel with an inlet and an outlet for holding a sample.
However, the prior art devices suffer from various shortcomings. One of these shortcomings is interference with biosensor readings caused by other substances in the sample fluid which can oxidize at the same potential. Prevalent among these are ascorbic acid, uric acid and acetaminophen. As these and other interfering substances oxidize, the current resulting from their oxidation is added to and indistinguishable from the current resulting from the oxidation of the blood analyte being measured. An error therefore results in the quantification of the blood analyte.
Another shortcoming is the interference caused by red blood cells (the hematocrit effect). This interference tends to cause an artificially high response rate for low hematocrit levels and, conversely, an artificially low response rate for high hematocrit levels.
Additional shortcomings of the prior art devices are that they have a more limited linear range and require a relatively large quantity of sample volume. Further, they require a relatively longer waiting time for development of a steady-state response before a reading can be achieved. Each of these shortcomings may, either individually or when combined with one or more of the other shortcomings, contribute to erroneous measurement readings during analysis. Preliminary tests performed by the inventors of the present invention have shown that the prior art which claims to reduce the effect of hematocrit on glucose readings, were limited to and worked only in lower glucose concentrations.
Because of the importance of obtaining accurate glucose readings, it would be highly desirable to develop a reliable and user-friendly electrochemical sensor which does not have all of the drawbacks mentioned above. Therefore what is needed is an electrochemical sensor that incorporates an interference-correcting electrode to minimize the interference caused by oxidizable substances present in the sample fluid. What is further needed is an electrochemical sensor whose response is substantially independent of the hematocrit of the sample fluid. What is still further needed is an electrochemical sensor which requires less sample volume than previously required by the prior art. Yet, what is still further needed is an electrochemical sensor which has a wide linear measurement range; that is, a sensor having a reduced or negligible interference effect and useable over a wider glucose concentration.